1. Field of the Invention
This invention pertains generally to chemical analytical and immunological testing, and particularly to processes wherein samples are analyzed by using self-operated mechanisms or devices, and more particularly to processes wherein a continuously flowing stream of a sample or carrier fluid is formed and flows into and through analysis wherein the continuously flowing stream is segmented by alternately injecting a sample, reagent or any number of fluids into a common flow path.
2. Description of the Related Art
There is a need for improved methods for measuring analytes in airborne particles. For example, there is a need for devices that can quickly detect and identify the presence of harmful materials in airborne infectious agents such as bacteria, fungi, or viruses, which transmit many diseases of humans, other animals, or plants. Some airborne proteins and pollens cause allergies. Improved methods for characterizing aerosols would be useful for understanding atmospheric chemistry, including the sources, chemical reactions, and fates of atmospheric particles.
Here, “airborne particle” refers to both the solid particles and liquid droplets in an air sample. The analyte is the specific molecule, microorganism, or virus to be identified. For example, for biological warfare (BW) agents that are protein toxins, e.g., ricin, the toxin itself is the analyte. For BW agents that are bacteria or viruses, the analyte can be a molecule that is specific to the bacteria or virus to be detected, e.g., a protein or a DNA or RNA sequence. In this case the amount of the analyte is measured; if this amount is significantly above a noise threshold, the presence of the BW agent is inferred. For BW agents that are bacteria or viruses, the analyte can be the bacteria or virus itself.
Objectives for some types of instruments needed for detecting BW-agents or other analytes in airborne particles are: sensitivity, specificity, rapid response, continuous operation, little need for consumables, little need for operator time, and be able to separate and store particles for further analysis.
Investigators have worked for years to develop instruments and methods that are useful for detecting airborne BW agents, and other harmful airborne particles. Samples can be collected from air using a variety of different collectors, and the collected samples can be subjected to many different types of microbiological and biochemical analyses.
Some reasons why it is so difficult for the above objectives to be met simultaneously are as follows. The analyte may comprise a small fraction of the mass of a highly complex sample. Sensitivity and specificity requires the sample to be mixed with one or more liquids, termed here, “analysis liquids,” where at least one of these liquids contains sensor molecules, also termed recognition molecules, that selectively bind to or interacts with the analyte. Example recognition molecules are antibodies and aptamers. Aptamers are DNA or RNA molecules that are selected for their ability to bind to the analyte. As a result of this binding of the recognition molecule to the analyte, some measurable property, e.g., fluorescence, must change according to the amount of analyte in the sample. That property is measured and the amount of analyte is inferred. Continuous operation requires continuous expenditure of consumables. Therefore, each measurement must require only a very small amount of consumables. In addition to the consumables used in analyzing the sample, consumables are typically expended in collecting particles from the air to be analyzed. If the particles are collected on filters or impacted on a surface, the filter or surface is a consumable unless it is cleaned, in which case whatever is used to clean it may be consumed. In typical analysis procedures for biochemical analytes in airborne particles, the airborne particles are collected into a liquid, which tends to evaporate as the sample is collected, especially if the air sample is warm and dry. Typically the consumable liquids, reagents, and/or filters, are sufficiently costly that it is too expensive to run instruments that identify biological molecules in air samples continuously for long periods of time.
The objectives of sensitivity and specificity, suggest choosing as analytes specific DNA sequences, and this approach may be applicable for some analytes. However, for a rapid response to spores, this approach is not feasible because 10's of minutes are required for the DNA from a spore to be extracted, amplified and detected. Also, this approach is not applicable to harmful substances that do not contain DNA or RNA, such as protein toxins.
A method to identify small amounts of analytes in airborne particles has been described, S. C. Hill, Aerosol Particle Analyzer for Measuring the Amount of Analyte in Airborne Particles, U.S. patent application Ser. No. 10/708,191, filed Feb. 14, 2004 and herein incorporated by reference. That application described a method for collecting particles into levitated droplets of an analysis liquid in order to rapidly detect analytes in the particles, and stated that the droplets of the analysis liquid could be collected and further analyzed. That invention was aimed specifically at the detection of analytes in particles without having the particles come into contact with any surface, so that there would be no problem of cross contamination from sample to sample. That application described the detection of the analyte by using a homogeneous assay in droplets levitated in a linear quadrupole. Manipulating multiple droplets levitated in a linear quadrupole was described by S. Arnold, P. Hendrie and B. V. Bronk, U.S. Pat. No. 5,532,140, Method and Apparatus for Suspending Microparticles, herein incorporated by reference. The method requires levitating or otherwise controlling the motion of a charged droplet for a sufficient time for the oppositely charged particles to combine with it.
H. B. Lin and S. C. Hill, Aerosol Particle Analyzer for Measuring an Analyte in Airborne Particles, U.S. patent application Ser. No. 10/816,579, filed Mar. 26, 2004, described an apparatus for collection of particles into a small volume of an analysis liquid at the end of a capillary tube, and where an optical change in the volume of liquid on the end of the capillary tube is detected if the analyte was in the particles that combined with that volume of analysis liquid. In that application, the rate of the collection of the particles is enhanced by electrostatic forces between the particles and the charged volume of liquid at the end of the capillary.
S. C. Hill and H. B. Lin, Aerosol Into Liquid Collector for Depositing Particles from a Large Volume of Gas into a Small Volume of Liquid, submitted to US Patent Office, Nov. 19, 2004, describes a means of collecting airborne particles into a small volume of liquid held at the end of a capillary.
For the two methods mentioned above in which the particles are collected into droplets held on the end of a capillary, there is some concern that particles may stick to the sides of the capillary.
Another potential problem for the methods described above, as far as measurement of biological analytes is concerned, is that the particles are given an electric charge, so that they have a charge that is opposite to that of the droplets and are attracted to the droplets. However, there is a concern that the charging of the particles may modify the biological analytes in these particles so that they can no longer be specifically analyzed.
Another limitation is as follows. The above methods collect all of the particles, having sizes above some minimum size, into the droplets for analysis (although not all with the same collection efficiency). Therefore, if, in an air sample, there is a very large excess of particles that do not contain analyte, then either (i) each droplet that contains a particle with analyte also contains many other particles that do not contain analyte, and therefore the sensitivity of the analysis may be reduced because of nonspecific binding problems, or (ii) a relatively large number of droplets is required for the analysis if there is to be only one or a very few particles per droplet to minimize problems from nonspecific binding, and so more of the instrument must be devoted to handling these droplets and measuring the fluorescence that occurs in them, and more of the reagents must be used, although for many cases the net use of reagents may be sufficiently small.